Date of Award

1988

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

First Advisor

R. J. Siebeling

Abstract

Species within the genus Vibrio can be identified serologically through detection of species specific H antigens expressed in the core protein of the polar flagella. Cholera vibrios also exhibit a specific cell wall polysaccharide antigen (A). Antibody reactive with these specific antigens can be employed for the rapid serological identification of Vibrio isolates. Species-specific anti-H sera were produced in rabbits immunized with flagellar core protein prepared from V. vulnificus. A coagglutination reagent was constructed by arming S. aureus Cowan I cells with the anti-V. vulnificus flagellar antibody. The reagent coagglutinated 99.3% of isolates identified bacteriologically as V. vulnificus and, other than V. pelagius, did not agglutinate 19 heterologous Vibrio species tested. In addition, monoclonal antibodies (Mabs) reactive with the flagellar core protein of either V. cholerae or V. vulnificus were produced and used to make coagglutination reagents. Staphylococcus aureus cells armed with Mab specific for V. cholerae flagellar core protein coagglutinated V. cholerae and V. mimicus isolates exclusively and did not react with 30 heterologous Vibrio species tested. These results suggest that V. cholerae and V. mimicus express similar H determinants. Latex beads armed with Mab reactive with V. vulnificus flagellar cores coagglutinated only V. vulnificus among 32 Vibrio species tested. The anti-H coagglutination tests represent a rapid, serologically specific, and inexpensive method for identifying V. cholerae, V. mimicus and V. vulnificus one step beyond primary isolation. Antibodies reactive with the A antigen of the cholera vibrios were employed in a membrane enzyme-linked immunosorbant assay (ELISA) to detect V. cholerae O1 isolates in enrichment cultures. The membrane ELISA detected V. cholerae Ogawa cells in 6 hour alkaline peptone (AP) enrichment cultures initially inoculated with approximately 10$\sp2$ colony forming units per ml. The detection system did not produce false positive results when overnight AP broth cultures inoculated with five heterologous Vibrio species were assayed. The membrane ELISA is a rapid and specific assay for the identification of O1 vibrios directly from enrichment cultures.

Pages

66

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